Bone is a complex, hierarchical composite consisting of both inorganic and organic components. This includes about 40-50% calcium deficient ion-substituted hydroxyapatite in the form of elongated crystals, about 30-40% type I collagen fibers, and the remaining 10% of water and cellular components. Osseous tissue is highly vascularized, dynamic, and adapts to changes in mechanical loading. Bone exhibits anisotropic behavior in that its mechanical properties are directionally dependent, and it resists loading best in the axial direction. Human compact bone has a compressive strength of about 170-200 MPa, a tensile strength of about 100-120 MPa, and a shear strength of about 60-80 MPa. Although bone is rather brittle, it does exhibit a certain degree of elasticity conferred by the type I collagen fiber reinforcement.
Calcium phosphate (CaP) bioceramics have been used for bone tissue engineering for decades. Calcium phosphates consist of three major elements: calcium, phosphorus, and oxygen, all of which are highly abundant on earth. Many calcium phosphates also include hydrogen as an acidic orthophosphate (i.e. H2PO4−) or as incorporated water (i.e. CaHPO4.H2O). Calcium phosphates are distinguished from each other by the type of phosphate anion in the complex and the oxidation state of the phosphorous. Calcium phosphates in particular contain the orthophosphate anion (PO43−) and include HA (hydroxyapatite) (Ca10(PO4)6(OH)2), alpha and beta TCP (tri-calcium phosphate) (Ca3(PO4)2), CDHA (calcium-deficient hydroxyapatite), MCPA (monocalcium phosphate anhydrous), MCPD (monocalcium phosphate dehydrate), DCPA (dicalcium phosphate anhydrous), DCPD (dicalcium phosphate dihydrate), TTCP (tetracalcium phosphate), and FA (fluoroapatite). Other materials that have been explored for their potential use as biomaterials for bone tissue engineering include bioglass and calcium silicate.
Although calcium phosphate ceramics are bioactive and biocompatible making them suitable for tissue engineering, there are several disadvantages of this material that prevent it from being used as an effective repair material for load-bearing bone defects. Specifically, while calcium phosphate ceramics exhibit high compressive strengths and can withstand large loading forces before fracture, they are extremely brittle. Brittle materials exhibit very low fracture toughness under loading, and have little to no impact resistance. Even though calcium phosphates, such as hydroxyapatite, closely mimic the chemical composition of the calcium phosphate material found in bone, CaP ceramics cannot match the fracture toughness of human bone (1.3 kJm−3) due to the lack of a tough, ductile component like collagen. Therefore, most research on calcium phosphate scaffolds for bone tissue engineering has focused on the use of these ceramics for non-load-bearing bone repair. Furthermore, the design of CaP bioceramic scaffolds generally requires a balance between porosity and mechanical strength. In order to encourage native bone ingrowth and osteogenesis within these ceramics, the scaffolds must exhibit a minimum degree of porosity (about 60-70% total pore volume) with pore sizes in the range of 50 to 100 microns. However, increasing the scaffold porosity of calcium phosphate ceramics significantly decreases the mechanical strength of the overall structure. Moreover, CaP ceramics degrade very slowly by a process of ion dissolution mediated by acidic environments. Thus, difficulties often arise in matching resorption rate with the rate of bone ingrowth within these materials.
A number of methods are known in the art for preparing calcium phosphate ceramic scaffolds and are described, for example, in Sylvain D., E. Saiz, A. Tomsia. Freeze casting of hydroxyapatite scaffolds for bone tissue engineering. Biomaterials. 2006. 27:32. 5480-89; Zang, Y., M. Zang. Synthesis and characterization of macroporous chitosan/calcium phosphate scaffolds for tissue engineering. Journal of Biomedical Research Materials. 2001. 55:3. 304-312; Zang, Y., M. Zang. Three-dimensional macroporous calcium phosphate bioceramics with nested chitosan sponges for load-bearing bone implants. Journal of Biomedical Research Materials. 2002. 61:1. 1-8; Varma, H. K., S. N. Kalkura, R. Sivakumar. Polymeric precursor route for the preparation of calcium phosphate compounds. Ceramics International. 1998. 24:6. 467-470; Soon, Y., K. Shin, Y. Koh, J. Lee, H. Kim. Compressive strength and processing of camphene-based freeze cast calcium phosphate scaffolds with aligned pores. Materials Letters. 2009. 63:17. 1548-50; Maccetta, A., I. G. Turner, C. R. Bowen. Fabrication of HA/TCP scaffolds with a graded and porous structure using a camphene-based freeze casting method. Acta Biomaterialia. 2009. 5:4. 1319-27; Potoczek, M., A. Zima, Z. Paszkiewicz, A. Slosarczyk. Manufacturing of highly porous calcium phosphate bioceramics via gel-casting using agarose. Ceramics International. 2009. 35:6. 2249-54; and Yang, T. Y., J. M. Lee, S. Y. Yoon, H. C. Park. Hydroxyapatite scaffolds processed using a TBA-based freeze-gel casting/polymer sponge technique. Journal of Materials Science: Materials in Medicine. 2010. 21:5. 1495-1502. Exemplary methods from the art include, but are not limited to:
Freeze-Drying: Calcium phosphate powders are mixed with water to create a ceramic slurry that is frozen and subsequently lyophilized to generate a freeze-dried ceramic green body with varying degrees of porosity depending on the ratio of ceramic powder to water. The resultant green body is then sintered.
Gel Casting: In situ polymerization of organic monomers in a ceramic slurry leads to rapid solidification and formation of ceramic green bodies. The organic gelling agents in the green bodies later act as sacrificial polymers during the sintering process to create voids in the final scaffold, imparting porosity in the finished product.
Dry Pressing: Dry calcium phosphate powder is packed and pressed into a mold to create a structurally stable blank and sintered to create the finished ceramic.
Sacrificial Porogens: Various synthetic polymer powders and inorganic porogens (i.e. naphthalene particles, PVC/PS/PEG/PMMA beads, cellulose acetate, resins, SiO2 particles, etc.) have been mixed with calcium phosphate ceramic paste to form green bodies. These porogens later act as sacrificial polymers by burning off during the sintering process thereby creating porosity in the finished scaffold. Other materials such as salts (NaCl, BaSO4, SrSO4) and liquids (camphene, oils) have also been used as porogens and binding agents for fabricating these calcium phosphate scaffolds.
Polyurethane Sponge Method: Porous polyurethane sponge is dipped into a calcium phosphate ceramic slurry and then sintered. The polyurethane sponge acts as a sacrificial polymer during the sintering process to create porosity in the finished ceramic scaffold.
Direct Foaming: Porous calcium phosphate ceramic materials are produced by introducing air bubbles into the CaP ceramic suspension. The resulting green state foams are subsequently sintered at high temperatures to obtain highly porous CaP ceramics.
However, the methods described above have several disadvantages that prevent them from generating CaP ceramics scaffolds that are sufficient for load-bearing orthopedic applications. First, these methods do not allow for the fabrication of highly stable calcium phosphate ceramic “green bodies”, defined as ceramic bodies that have not yet been sintered or fired. This prevents the fabrication of complex geometric shapes, and limits scaffolding to simple geometries (cylinders, rectangles, etc.). The ability to generate CaP ceramic scaffolds of complex geometry would allow for patient-specific design of ceramic bone grafts to fit the particular defect geometry. In addition, the methods of CaP scaffold fabrication mentioned above do not allow for the creation of high strength CaP scaffolds that can match the mechanical properties of human cortical bone. This is mainly due a lack of control over the total porosity and pore size in the finished scaffold.
Thus, there is a need in the art for methods of fabrication of high strength, complex geometry calcium phosphate ceramic scaffolds formed from highly stable calcium phosphate ceramic green bodies. Additionally, there is a need in the art for methods of fabrication of high strength calcium phosphate ceramic scaffolds matching that of human cortical bone.